Essential role of MYB transcription factor: PvPHR1 and microRNA: PvmiR399 in phosphorus‐deficiency signalling in common bean roots

Abstract

Phosphorus (P), an essential element for plants, is one of the most limiting nutrients for plant growth. A few transcription factor (TF) genes involved in P-starvation signalling have been characterized for Arabidopsis thaliana and rice. Crop production of common bean (Phaseolus vulgaris L.), the most important legume for human consumption, is often limited by low P in the soil. Despite its agronomic importance, nothing is known about transcriptional regulation in P-deficient bean plants. We functionally characterized the P-deficiency-induced MYB TF TC3604 (Dana Farber Cancer Institute, Common Bean Gene Index v.2.0), ortholog to AtPHR1 (PvPHR1). For its study, we applied RNAi technology in bean composite plants. PvPHR1 is a positive regulator of genes implicated in P transport, remobilization and homeostasis. Although there are no reports on the regulatory roles of microRNAs (miRNA) in bean, we demonstrated that PvmiR399 is an essential component of the PvPHR1 signalling pathway. The analysis of DICERlike1 (PvDCL1) silenced bean composite plants suppressed for accumulation of PvmiR399 and other miRNAs suggested that miR399 is a negative regulator of the ubiquitin E2 conjugase: PvPHO2 expression. Our results set the basis for understanding the signalling for P-starvation responses in common bean and may contribute to crop improvement.

An Autophagy-Related Kinase Is Essential for the Symbiotic Relationship between Phaseolus vulgaris and Both Rhizobia and Arbuscular Mycorrhizal Fungi

“…To create the PvPI3K-RNAi construct, a 537-bp fragment corresponding to the 39-coding region of Pv-PI3K was amplified by PCR using the primers PvPI3K-RNAi_F and PvPI3K-RNAi_R (Supplemental Table 1) and cDNA from P. vulgaris nodules (20 dai). After cloning this fragment into pENTR/D-TOPO (Invitrogen) and confirming the presence of the insert by sequencing, the PCR product was recombined into the compatible recombination sites of the Gateway-based hairpin pTDT-DC-RNAi vector (Valdés-López et al, 2008). The correct orientation of the recombined fragments was confirmed by sequencing and by PCR using the primers Wrky-3_F or Wrky-5_R with PvPI3K-RNAi_F (Supplemental Table 1), as described (Valdés-López et al, 2008).…”

“…A specific PvPI3K-RNAi construct (537 bp) was cloned downstream of the 35S promoter sequence in the Gateway-based hairpin vector pTdT-DC-RNAi (Valdés-López et al, 2008) to generate pTdT-PvPI3K-RNAi. A previously described construct bearing a truncated and irrelevant sequence from Arabidopsis pre-mir159 cloned in pTdT-DC-RNAi (Montiel et al, 2012) was used as a control vector and is herein referred to as pTdT-Sac-RNAi.…”

Validating Genome-Wide Association Candidates Controlling Quantitative Variation in Nodulation

“…The transformations were conducted using a modified version of the binary vector backbone pNB96 (pSC218; Curtin et al, 2011) with an introduced inverted-repeat cassette to facilitate cloning of hairpin RNA fragments (pSC218GH) and a reading-frame cassette to incorporate both the Cas9 and target guide RNA entry vectors (pSC218GG). The inverted-repeat and reading-frame cassettes were derived from previously reported pTDT-DC-RNAi and pTDTO vectors (Valdés-López et al, 2008) and were driven by the constitutively expressed Glycine max Ubiquitin promoter (Hernandez-Garcia et al, 2010).…”

RNA interference: concept to reality in crop improvement

“…PvPHR1 is supposed to be positive regulator of genes implicated in transport, mobilization and homeostasis of phosphorus. Valdés-López et al (2008) demonstrated that miRNA399 of P. vulgaris (PvmiR399) is an essential component of the PvPHR1 signalling pathway in common bean (P. vulgaris L.). Jones-Rhoades and Bartel (2004) have indicated that miR395 and miR399 are up-regulated when plants suffer from sulphur and phosphate starvation in Arabidopsis.…”